Normal Tissue Injury

Collateral normal tissue injury during radiotherapy is common despite technological advances in radiation dose delivery and distribution within the treatment volume. Moderate dose exposures (up to 10-Gy) may also occur from radiation accidents or terrorism, leading to symptoms that may appear a few weeks, months, or years later. Therefore, the development of biomarkers and radiation effect modulators with mechanistic underpinnings are essential to improve outcomes for cancer patients and individuals accidentally exposed to radiation.Reference Coleman, Blakely and Fike ¹ The risk and severity of normal tissue reactions are dose-dependent, tissue-specific, and vary with time after treatment. Thus, treatment approaches to reduce the severity of normal tissue injury should consider these factors to improve the patient’s quality of life.Reference Stone, Coleman and Anscher ²

Building on the strengths of multiple systems for grading the adverse effects (AEs) of cancer treatment, Common Terminology Criteria for Adverse Events (CTCAE) 3.0 guidelines were developed. Seminal contributions of Dr. Coleman to CTCAE 3.0 involved building consensus among the oncology research community, the first comprehensive, multimodality grading system consisting of acute and late effects, including radiotherapy. With subsequent multiple iterations, this work on CTCAE now serves as the standardized vital tool for reporting AEs in clinical trials.Reference Trotti, Colevas and Setser ³

Bringing research advances in the development of radiation countermeasures for mass casualties in the face of the September 11, 2001 terrorist attack, and with the continuing goal to reduce normal tissue injury from radiotherapy, required understanding the impact of protectors and mitigators on tumor and normal tissue response. Advances in tissue biology, injury mechanisms, predictive biomarkers, and radioprotection have revitalized normal tissue protection efforts. A pipeline of drugsReference Prasanna, Narayanan and Hallett ⁴ and biomarkersReference Prasanna, Narayanan and Zhang ⁵ is under development in this space. Key adverse effects of radiation and drug combinations were reviewed, highlighting current limitations in radioprotection strategies and proposing new concepts for improving radiation therapy.Reference Prasanna, Stone and Wong ⁶

Radiation-induced pulmonary fibrosis (RIPF), a progressive, irreversible, and clinically significant intermediate to a late-occurring side effect, is a consequence of normal tissue injury. While pre-clinical models show that various molecular targets can be exploited to prevent or mitigate RIPF, further basic and translational research is needed to identify optimal therapies and practical biomarkers for prediction.Reference Citrin, Prasanna and Walker ⁷ Inhibiting the conversion of Angiotensin Converting Enzyme (ACE) ACE with drugs such as Captopril and Losartan can ameliorate radiation fibrosis in various tissues.Reference Moulder, Cohen and Fish ⁸ Dr. Coleman’s lab’s recent analysis of the published literature on RIPF, miRNA, and therapy-induced senescence (TIS) suggested RIPF is regulated through several miRNAs, paving the way for the potential discovery and development of miRNA therapeutics and the development of early biomarker panels to predict RIPF.Reference Prasanna, Aryankalayil and Citrin ⁹

Collateral normal tissue injury from radiotherapy is unavoidable, with symptoms potentially emerging weeks to years post-treatment; thus, developing biomarkers and radiation effect modulators is crucial for improving patient outcomes. Advances in radiation countermeasures and understanding the mechanisms of injury are essential to mitigate radiation-induced effects, such as pulmonary fibrosis. At the same time, continued research focus is crucial to identify effective therapeutic strategies and predictive biomarkers.

Radiation Immunotherapy Combinations

Cancer immunotherapy is emerging as an approach for treating cancer patients. However, several critical challenges persist in this vital field. Key issues include identifying the most effective combinations of therapies and the difficulties in translating preclinical findings into successful patient treatments.Reference Hegde and Chen ¹⁰ Exploring the fundamental aspects of T-cell-tumor interactions is crucial for enhancing immunotherapy combinations, especially in conjunction with external beam radiotherapy for solid tumors. Radiotherapy can influence the lytic efficiency of CD8+ cytotoxic T lymphocytes (CTLs). In a pioneering study using a mouse adenocarcinoma model, Dr. Coleman’s team, in collaboration with James Hodge, demonstrated that irradiation is critical in regulating Fas pathways and sensitizing tumors to antigen-specific CTLs. This research suggests combining tumor-targeted irradiation with CTLs can lead to more effective antitumor responses.Reference Chakraborty, Abrams and Camphausen ¹¹

Further, immune checkpoint inhibitors (ICIs) can also effectively treat cancers with pre-existing immune responses, particularly in “hot” tumors with significant lymphocytic infiltration. In contrast, “cold” tumors, lacking this infiltration, typically resist immunotherapy. Preclinical studies suggest radiotherapy may enhance ICI efficacy by recruiting T cells into cold tumors. However, to advance this approach, rigorous trials are needed to address critical questions, such as defining the optimal radiation doses, sequencing with ICIs, the effect of metastasis location, and the durability of T-cell responses induced by this combination.Reference Demaria, Coleman and Formenti ¹² Radiotherapy may enhance immunotherapy responses by inducing the DNA exonuclease Trex1, which limits the immunogenicity of cancer cells by degrading cytosolic DNA generated from radiation. By optimizing radiation doses that avoid Trex1 induction, increased interferon-β production can recruit dendritic cells and prime CD8+ T cells, leading to systemic tumor rejection in combination with immune checkpoint blockade. Thus, Trex1 induction may guide the selection of radiation dose and fractionation in patients treated with immunotherapy.Reference Vanpouille-Box, Alard and Aryankalayil ¹³

Some evidence suggests that radiotherapy can enhance immune responses to tumors, particularly advanced delivery techniques such as hypofractionation. However, there is still limited understanding of how different radiation fractionation regimens impact immune modulation, necessitating further studies to optimize radiotherapy-immunotherapy combinations. A consensus discussion among radiation oncologists and immunologists highlighted the need for tailored clinical study designs that account for variations in radiation dose- and fractionation and the need for immune biomarkers to better evaluate the effects of these therapies.Reference Demaria, Guha and Schoenfeld ¹⁴ Numerous randomized clinical trials of radiotherapy–immunotherapy combinations have not shown significant therapeutic benefits compared to each treatment alone. This may be due to suboptimal study design and standard radiotherapy schedules that do not consider immunostimulatory effects. To enhance the anti-tumor immune response and achieve meaningful clinical benefits, successful combinations may require modifications to standard radiotherapy regimens and target volumes, allowing for better preservation of immune function while maximizing cancer cell killing.Reference Galluzzi, Aryankalayil and Coleman ¹⁵

Cellular senescence acts as a tumor suppressor, preventing the spread of damaged cells, and is a mechanism through which cancer therapies may also work. However, radiation and chemotherapy can lead to an accumulation of senescent cells, which may paradoxically enhance tumor relapse, metastasis, and treatment resistance via the senescence-associated secretory phenotype (SASP). A workshopReference Prasanna, Citrin and Hildesheim ¹⁶ envisioned by Dr. Coleman discussed senescence research, therapy-induced heterogeneity, and a “one-two punch” strategy for combining senescence induction followed by a selective clearance of senescent cells, highlighting the knowledge gaps and future directions for improving cancer treatment outcomes. The context of senescence in tumor and normal tissue was outlined in detail with discussion focusing on toxicity (normal tissue) and radioresistance (tumor) mitigation, areas upon which Dr. Coleman wished to focus.Reference Prasanna, Citrin and Hildesheim ¹⁶

Considering the limitations and complexities of the cell-killing-based cancer treatment approaches, a provocative approach to combining immunotherapy with cytotoxic drugsReference Marin, Boix and Garcia-Garijo ¹⁷ or radiotherapy might include inducing senescence specifically in the tumor microenvironment to trigger a robust immune response, followed by using senolytics to eliminate senescent cells to prevent tumor maladaptation and adverse effects in normal tissues, and unleash long-lasting antitumor immunity with immunomodulators.Reference Prasanna ¹⁸ However, such an approach will require scientifically rigorous “fit-for-purpose,” well-controlled preclinical research and development involving dose and schedule optimization of radiation and drugs, using representative in vitro and in vivo cancer models to obtain high-quality data to proceed to clinical studies.Reference Prasanna ¹⁸

Cancer immunotherapy is gaining traction as a treatment option, yet challenges remain. Evidence suggests combining radiotherapy with immunotherapy, particularly in targeting tumor microenvironments and optimizing treatment regimens, could enhance immune responses and improve patient outcomes. However, further studies are needed to refine these strategies and address the complexities of cellular senescence and immune modulation.

Rigor and Reproducibility of Preclinical Data

Successful translation of preclinical laboratory data to patient treatments through clinical trials requires high-quality, reproducible data with robust predictive value. There is an urgent need to enhance the reproducibility and translatability of preclinical findings, especially for molecular therapeutics involving drug-radiation combinations, such as combinations with immunotherapy. Current assays, like the clonogenic assay, are vital for in vitro research.Reference Besch, Tanner and Sattler ^{19 –}Reference Franken, Rodermond, Stap and Haveman ²¹ Still, advanced methods—including 3D culture models, organoids, tissue chips, and in vivo studies with genetically engineered mouse models—can provide deeper insights into treatment mechanisms and responses to therapy. To achieve reliable outcomes, standardization of procedures and careful selection of endpoints are crucial. Additionally, incorporating radiation modifiers into standard clinical care is necessary for thoroughly evaluating their effects in combination therapies.Reference Coleman, Higgins and Brown ²²

Drug–radiation combinations can enhance clinical outcomes, but most studies use empirical methods without considering mechanistic synergies in cell death or the interconnectivity of molecular pathways. This oversight often leads to suboptimal efficacy and increased toxicity. However, there are opportunities to strengthen preclinical evidence through “fit-for-purpose” translational studies that simulate clinical scenarios. Utilizing standardized assays and algorithms to evaluate molecular interactions can inform the selection of dose, fractionation, and scheduling. Critical issues in translating preclinical data to improve the relevance of assays like in vitro clonogenic survival, apoptosis, in vivo tumor regression, and organ toxicity evaluations while fostering innovation in emerging areas of preclinical radiobiology are recently discussed.Reference Prasanna, Ahmed and Hong ²³

Dr. Coleman was a strong advocate of the successful translation of preclinical laboratory data to patient treatments through clinical trials, which necessitates high-quality, reproducible data, emphasizing the need for advanced methodologies and standardization to enhance the efficacy and safety of drug-radiation combinations, particularly in the context of molecular therapeutics and immunotherapy.

RNA Biomarkers in Biodosimetry: Key Indicators of Radiation-Induced Damage

Dr. Coleman’s research at the National Cancer Institute for the last 24 years has yielded significant scientific advancements, as evidenced by his laboratory’s high-impact publications. Key achievements include discovering radiation-inducible targets that rapidly change during treatment and exhibit long-term persistence.Reference Coleman, Eke and Makinde ^{24 –}Reference Eke, Aryankalayil and Bylicky ²⁷ Using multiple cell lines and tumor models, the Coleman lab demonstrated that these targets can be effectively treated with drugs that previously had limited or no efficacy before radiation therapy. Additionally, these findings suggest that current radiation and combined modality treatment strategies, which involve multiple cycles, may contribute to the challenges in cancer treatment. However, this research has also paved the way for new explorations, such as immune modulationReference Vanpouille-Box, Alard and Aryankalayil ^{13 ,} Reference Galluzzi, Aryankalayil and Coleman ¹⁵ and normal tissue injury at clinically relevant doses. He made significant contributions to the field of biodosimetry by helping establish NIAID’s research and development programs focused on radiation injury resulting from whole or partial body exposure following a nuclear detonation.

He recognized the urgent need for rapid and reliable biological dosimetry methods, especially compared to standard biodosimetry tools like dicentric chromosome aberration (DCA) analysis performed using peripheral blood lymphocytes obtained from individuals exposed to radiation, to ensure effective clinical management and resource allocation during large-scale nuclear disasters.Reference Prasanna, Moroni and Pellmar ^{28 –}Reference Sullivan, Prasanna and Grace ³⁰ We developed RNA-based biomarkers to predict radiation-induced damage based on the laboratory’s previous research into radiation-induced endothelial cell injury and our expertise in RNA biology from radiation fractionation studies.Reference Chopra, Shankavaram and Bylicky ^{31 –}Reference John-Aryankalayil, Palayoor and Makinde ³³ Additionally, the lab leveraged innovative “organ-on-a-chip” models to enhance understanding of organ-specific markers following radiation injury to identify potential mitigators to reduce radiation therapy-induced damage.Reference Martello, Ueda and Bylicky ³⁴

To identify dose-differentiating microRNA (miRNA) signatures, the Coleman lab used in vitro cell culture models in addition to in vivo mouse, mini-pig, and non-human primate (NHP) models after whole- and partial-body radiation.Reference Chopra, Shankavaram and Bylicky ^{31 ,} Reference Aryankalayil, Chopra and Levin ³⁵ From the whole-body irradiated mouse studies, the lab observed a time- and dose-responsive alteration in the expression levels of miRNAs. A substantial number of the miRNAs were altered at 24- and 48-hour time points post-irradiation. The lab identified a 23-miRNA signature using feature selection algorithms and classifier models. An inverse correlation in the expression level changes of miR-17 members and their targets were observed in whole-body irradiated mice and non-human primates.Reference Aryankalayil, Chopra and Makinde ³⁶

Considering the pivotal role of endothelial cells in driving radiation-induced organ injury, he recognized their significance in the biomarker signatures observed in circulation. Based on our previous work showing the endothelial cells’ dose and time response to single doses of radiation (1 Gy up to 10-12 Gy), these cells could, therefore, provide biomarkers of response and insight into mechanisms of the radiation response. Organ-specific RNA biomarkers for liver, lung, and heart damage are being analyzed to potentially guide treatment and develop targeted medical countermeasures for use in disaster triage and the clinic.Reference Aryankalayil, Martello and Bylicky ^{37 –}Reference Aryankalayil, Bylicky and Martello ³⁹ Recently, the lab used a liver-on-a-chip model to validate several of the biomarkers we identified in whole-body irradiated mice. This model mimics the complexity of the organ as the chips can be seeded with the 4 major liver cell populations: hepatocytes, Kupffer cells, stellate cells, and liver sinusoidal endothelial cells (LSECs). This model offers several advantages. In addition to reducing the need for animal studies to enhance biological relevance and improve translation, the chip platform provides the advantage of studying liver-specific cell interactions without the confounding effects of systemic variables, which are difficult to control in live animal models.

These are best suited to perform initial drug screenings and establish optimal and most effective concentrations.

The Coleman lab used RNA biomarkers (mRNA, miRNA, and lncRNA) to predict survival and understand how radiation damages tissue to develop targeted mitigators and treatments to address radiation exposure and normal tissue injury using an integrated RNA biomarker approach using a combinatorial approach of using different RNA biomarkers instead of depending on a single marker.Reference Coleman, Eke and Makinde ²⁴ He envisioned using a decision tree model to predict radiation exposure in a mass causality nuclear incident (Figure 3B). The lab’s expertise in RNA biology also allowed them to investigate coding and non-coding RNAs as biomarkers of cell adaptation for use as therapeutic targets.Reference Eke, Bylicky and Sandfort ⁴⁰ The omics data showed stable upregulation of lncRNA LINC00261 and LINC00665 after 2 months of radiotherapy, which was more pronounced after SD irradiation. Notably, patients with a high expression of LIN00261 have a worse prognosis. Knockdown of LINC00261 and LINC00665 in long-term PC3 cells decreased survival after re-irradiation and affected DNA double-strand break repair. Mechanistically, both lncRNAs showed an interdependent and regulated expression of the DNA repair proteins CtIP (RBBP8), XPC, and the microRNA miR-329.

Under his leadership, the lab’s research focused on utilizing RNA markers for 3 critical applications, as shown in (Figure 2). First, for radiation oncology applications to investigate radiation-inducible changes for potential molecular and immunological targeting. This included basic adaptation mechanisms to clinically relevant radiation treatments supported by the Center for Cancer Research at the National Cancer Institute (NCI). Second, his work employed RNA biology for mass casualty triage and patient management. Biomarkers of the whole body or extensive partial body radiation exposure were identified using mice, human organ-on-a-chip, NHP, and minipig models, focusing on early time point markers. With these biomarkers, he was particularly interested in developing point-of-care diagnostics and radiation sensitizers or mitigators. Third, he was enthusiastic about identifying organ-specific RNA markers of injury in patients undergoing radiation therapy. These markers, detectable in circulation at an early stage, could predict the late effects of radiation therapy and ultimately enhance the quality of life for cancer patients.

Figure 2.

RNA markers in radiation biology: applications for diagnosis and treatment.

Using RNA markers as a panel of assays has multiple practical applications in the radiation field and will be key in predicting required therapies and optimizing radiation injury management.

These findings underscore the urgent need for advancements in rapid diagnostic technologies and mitigation strategies following radiation exposure. The early establishment of whole-body radiation response within 24 hours highlights the importance of timely intervention in preventing late effects of radiation damage. Developing RNA markers for swift and accurate assessment can significantly enhance triage capabilities and inform targeted medical care for affected individuals, especially when clinical symptoms may not yet be apparent (Figure 3). By utilizing gene expression and non-coding RNA profiling, one can identify at-risk patients and implement interventions before severe organ injuries occur. Additionally, while current biodosimetry assays provide valuable insights into physical doses, a more precise approach to assessing specific organ injuries could greatly improve patient outcomes. Integrating a comprehensive panel of assays (Figure 3) will be crucial in predicting necessary therapies, ultimately leading to better management of radiation injuries. Continued research in this area is vital to translate these findings into practical applications that can save lives and reduce the long-term consequences of radiation exposure.

Figure 3.

Methodology for utilizing RNA markers in liquid biopsy for dose prediction and cancer treatment. A. Schematic overview of the approach for predicting radiation exposure and tissue damage using RNA-based signatures derived from liquid biopsy. Workflow beginning with a minimally invasive liquid biopsy (blood draw or urine sample), followed by the use of high-throughput diagnostic platforms to identify RNA markers. Multiple RNA biomarkers on a lab on a chip model will be highly useful in case of mass triaging radiation exposure scenario. B. Application of predicted RNA signatures for triaging individuals exposed to accidental radiation and evaluating normal tissue injury in patients undergoing radiotherapy.

Radiation Research Program Initiatives Including Health Disparities

In addition to normal tissue biology, immune-oncology, and RNA biomarkers development taking place within his laboratory or his collaborators’, Dr. Coleman directed the national research community via his role in supporting extramural science toward other advanced research areas, particularly the need to link physics and radiobiology.Reference Griffin, Ahmed and Amendola ^{41 ,} Reference Buchsbaum, Jaffray and Ba ⁴² The promising domain of particle therapy exemplified this. Working with members of RRP, he advocated for studies that examined the efficacy and biological implications of particle therapy, seeking to understand its potentially unique effects on cancerous and healthy tissue, ultimately helping the field via set-aside funding. ⁴³ These and subsequent efforts in the extramural division, such as the Preclinical Chemoradiotherapy Testing Consortium,Reference Buchsbaum, Coleman and Bernhard ⁴⁴ prioritized cutting-edge research and ensured that the findings could be translated into clinical contexts.

Dr. Coleman also actively engaged the underrepresented communities, exemplified by his work with the RRP team co-led by Dr. Rosemary Wong to include the Native American community in radiation research initiatives. This focus highlighted NCI’s commitment to diversity and inclusivity in science and acknowledged the need to address health disparities in radiation therapy research. The collaboration aimed to involve Native American scientists and communities in radiation oncology studies, laying the groundwork for increased representation in research and health care.Reference Petereit, Rogers and Burhansstipanov ^{45 ,} Reference Coleman, Wong and Petereit ⁴⁶

In the final years of his NCI extramural leadership, Dr. Coleman shifted his focus to the creation of a foundational molecular radiation biology consortium to address very complex normal tissue and tumor tissue dynamic, heterogeneous biology during radiation oncology working closely on the science with Dr. Jeff Buchsbaum in the creation of the Radiation Oncology Biology Integration Network grant program.Reference Buchsbaum ^{47 – 51} This consortium investigates the foundational molecular biology underlying radiation therapy. It emphasizes the importance of targeting specific molecular pathways to improve the precision of radiotherapy and the concept of optimal biologic dose.Reference Buchsbaum, Espey and Obcemea ^{48 ,} Reference Ahmed, Coleman and Mendonca ⁵² This ongoing work will hopefully underscore the value of understanding cellular responses at the molecular level and of workforce development, helping establish radiation therapy as a more exact science and enhancing its effectiveness for patients worldwide.

Beyond his scientific contributions, Dr. Coleman has had an enduring impact on global health policy and radiation oncology through his dedication to promoting equitable access to cancer care worldwide.Reference Abdel-Wahab, Coleman and Eriksen ⁵³ In an interview with the American Cancer Society shortly before he passed away, Dr. Coleman said, “I hope that 25 years from now, everyone around the world will have access to some reasonable level of the full spectrum of cancer care.”Reference Printz ⁵⁴ He was instrumental in founding the International Cancer Expert Corps (ICEC), a non-profit organization dedicated to expanding cancer treatment capacity in low- and middle-income countries.Reference Coleman, Formenti and Williams ⁵⁵ ICEC has bridged gaps in global cancer care, providing a platform for health care professionals to train and mentor others in underserved regions. Although ICEC’s work and Dr. Coleman’s role in its development are discussed in detail elsewhere in this special issue, his founding vision for the organization underscores his lifelong commitment to global health and social responsibility that impacted his vision for his work in NCI at all levels.

Dr. Coleman’s work with the International Atomic Energy Agency (IAEA) and partnerships with various countries further exemplified his dedication to extending the benefits of radiation therapy beyond US borders. Collaborating with IAEA and other international agencies, he advocated for initiatives that promoted radiation oncology and bolstered global health security through improved health care infrastructure and radiation safety standards. These efforts aimed to ensure that safe and effective radiation treatment options would be available to diverse populations worldwide, contributing to improved cancer care and broader health outcomes globally.Reference Ige, Jenkins and Burt ^{56 –}Reference Coleman, Pomper and Chao ⁶¹

As a senior leader within the NIH, Dr. Coleman’s influence extended to both his peers and the next generation of scientists. His mentoring junior scientists and physicians has shaped numerous careers in radiation oncology and related fields. Dr. Coleman’s guidance fostered a collaborative spirit within NIH and beyond, emphasizing the importance of interdisciplinary efforts and shared goals in scientific pursuits. Many of his mentees, now established leaders, continue to advance the fields of radiation biology and oncology, carrying forward his legacy of dedication, integrity, and commitment to improving patient outcomes.Reference Coleman and Love ^{62 –}Reference Vapiwala, Thomas and Grover ⁶⁴

Norm came into work early, often to take international and national calls of significance, and always wanted to discuss new articles across many disciplines. In this capacity, he could meet and discuss science with each member of RRP as an expert, as well as most of the other areas of NCI. He took great pride in reading background data of the day’s meeting topics in depth to render fair decisions on what to fund and support. Above all else, he focused on being kind, just, and truthful. To him, integrity was an absolute quality to be upheld, maintained, and respected. He actively listened and was present for those in RRP no matter what else was happening in his world.

Summary

Dr. Coleman’s contributions to radiation oncology through his intramural and extramural leadership roles at NCI have been transformative. His pioneering work in radiation biology research, including normal tissue injury, radiation-immunotherapy combinations, improving the predictive value of preclinical radiobiology research to enhance translation and disaster management science and new areas of RNA biomarkers and tissue chips, have long-lasting impact. His management of NCI’s RRP propelled the field forward, from establishing research standards to championing optimal, biological dose, radiation molecular biology integration, and inducible targets. His legacy extends internationally through his mentorship, his co-founding International Cancer Expert Corps (ICEC), and his many collaborations, such as those with the IAEA, promoting equitable access to cancer care and enhancing global health security. Dr. Coleman’s influence as a scientist, teacher, public servant, mentor, and visionary will continue to resonate within the scientific community for years to come. His kindness and support helped so many inside and outside of NCI.